The neurotoxins produced by Clostridium botulinum are among the most potent toxins known, leading to life threatening acute flaccid paralysis. Current measures for treating botulism are all limited to macromolecular antitoxins, which must be administered at the onset of the disease to prevent long-term damage or death. Hence, there is a great need for cell-permeable small molecules that inhibit these bacterial metalloproteinases, which may find use as biological weapons. The goal of this proposal is to identify small molecular weight BoNT inhibitors with broad spectrum activity against the various subtypes, and possibly even the various serotypes. Our structure-guided combinatorial chemistry approach encompasses the following key features: (a) Structure-based design of combinatorial libraries of'divalent' molecules that may bind to the enzyme's active site through a substrate-like fragment and a metal-coordinating group, (b) designing substrate mimics by replacing peptide bonds with click chemistry-derived linkages, and (c) applying medicinal chemistry principles to increase the chances for improved bioavailability. The specific aims are as follows: (1) Design and produce several hydroxamate-based libraries that are targeted against BoNT metalloproteinases; (2) Design and produce several libraries of peptidomimetic compounds that contain heterocyclic Zinc ligands; (3) Develop predictive computational tools for binding-affinity and targetspecificity trends.